The present invention relates to methods of treating cancer using antibodies directed against p53 mutant proteins.
The tumor suppressor gene p53 inhibits tumor growth primarily via induction of apoptosis. Mutations in the p53 tumor suppressor gene are the most common genetic alterations and occur in more than half of all human tumors. Approximately 90% of these alterations are missense mutations in the DNA-binding core domain responsible for sequence-specific binding of wild-type p53 protein to target genes. Many of these mutations cause a common conformational change in the p53 protein, which results in the exposure of an epitope that is otherwise hidden inside the wild type p53 molecule.
The involvement of p53 mutants in cancer progression was suggested to be associated either with trans-dominant suppression of wild-type p53 or a wild-type p53-independent oncogenic “gain of function”. As wild-type p53 forms a tetramer to exert its tumor suppressor activity, it is generally accepted that heteromerization of mutant p53 with wild-type p53 drives the wild-type protein into a mutant or otherwise inactive conformation which leads to the trans-dominant suppression phenomenon. The “gain of function” of mutant p53 may be attributed to two mutually possible mechanisms. One is the abrogation of the tumor suppressor activity of p53 family members, p63 and p73, that were found to physically interact with mutant p53, but not with the wild-type p53 protein, and to interfere with their activity. The second involves the ability of mutant p53 to trans-activate or repress specific genes that mediate the various oncogenic activities of these mutants. Core domain p53 mutants were found to trans-activate genes, such as multiple drug resistance (MDR-1), c-myc, proliferating cell nuclear antigen (PCNA), interleukin-6 (IL-6) and epidermal growth factor receptor (EGFR) and early growth receptor (EGR-1), these genes being different from those reactivated by wild type p53.
Given the active role of p53 mutants in promoting tumorigenicity, efforts have been made to inactivate their function or to revert them into a wild-type phenotype. These include the introduction of second site suppressor mutations (e.g., N239Y, N268D and H168R) that can at least partially restore specific DNA binding to mutant p53. In addition, synthetic peptides derived from the C-terminus of the p53 protein, or the CDB3, a p53-binding protein (p53BP2) derived compound, were found to restore DNA binding followed by transcriptional trans-activation, as well as induction of p53 dependent apoptosis to tumor cells. Moreover, low molecular weight compounds, such as CP-31398 and PRIMA-1, were shown to restore wild type conformation, transcriptional trans-activation and to induce apoptosis in cells and in human tumor xenografts carrying mutant p53. However, such peptides and compounds lack the ability to distinguish between the wild-type and mutant form of p53, a property crucial for targeted treatment.
Thus, novel anti cancer treatment modalities which specifically target a broad range of p53 mutants and not wild-type p53 proteins are desired
More than 90% of the mutations found in the p53 protein produce a conformational change in the p53 protein which results in the exposure of an epitope, which is otherwise hidden in the hydrophobic core of the molecule. Such an epitope was localized to amino acids 212-217 of the human p53 protein (GenBank Accession No. NP—000537) or amino acids 209-214 of the mouse p53 protein (GenBank Accession No. NP—035770) and has a sequence of FRHSVV (SEQ ID NO:1). Prior studies describe the isolation of a single-chain scFv mouse antibody prepared from a mouse immunized with SEQ ID NO:1. This antibody (named ME1), was found to be expressed in the cytosol of mammalian cells and to bind mutant p53 protein but not the wild-type p53 protein with an affinity of 10−7 M (Govorko D, Cohen G and Solomon B., 2001, J. Immunol. Methods. 258: 169-81). However, although this antibody presents a useful tool for clarifying the role of mutant p53 in tumor transformation, due to its mouse origin and its moderate affinity towards the mutated p53, its therapeutic application is limited.
There is thus a widely recognized need for, and it would be highly advantageous to have, a human derived antibody capable of specifically targeting with high affinity mutant p53 proteins.